Compositions and methods comprising prostaglandin related compounds and trefoil factor family peptides for the treatment of glaucoma with reduced hyperemia

ABSTRACT

Compositions, methods, and pharmaceutical products related to prostaglandin-related compounds and trefoil factor family peptides are disclosed herein. Of particular interest are compositions and methods useful for the treatment of glaucoma with a reduced occurrence of hyperemia.

CROSS REFERENCE TO RELATED APPLICATIONS

This is a national stage application under 35 U.S.C. § 371 of PCTapplication PCT/US2004/02777, filed on Aug. 25, 2004, which claims thebenefit of Provisional Application No. 60/508,445, filed on Oct. 3,2003.

FIELD OF THE INVENTION

The present invention relates to pharmaceutical compositions comprisingprostaglandin-related compounds and trefoil factor family peptides.

BACKGROUND OF THE INVENTION Description of Related Art

Active drugs often have undesirable side effects at theirtherapeutically effective concentrations. This is particularlyproblematic for topical use in sensitive areas such as the eyes, whereirritation is very difficult to avoid even for relatively mild drugs. Asa result, formulating topical ophthalmic drugs is a particularlychallenging problem. This is unfortunate because topical ophthalmic useof drugs has been found to be very useful in managing many conditionsaffecting the eye such as dry eye, infection, inflammation, allergy, andglaucoma. Glaucoma is a particularly devastating disease of the eyecharacterized by increased intraocular pressure, which is often treatedby topical ophthalmic application of a drug. Glaucoma occurs in about 2%of all persons over the age of 40 and may be asymptotic for years beforeprogressing to rapid loss of vision. In cases where surgery is notindicated, many drugs have been found to be useful in treating glaucomaby topical application including β-adrenoreceptor antagonists andα₂-adrenoreceptor agonists. Recently, prostaglandins have been shown tobe particularly useful in the topical treatment of glaucoma.

Whereas prostaglandins appear to be devoid of significant intraocularside effects, ocular surface (conjunctival) hyperemia, foreign-bodysensation, and itching (pruritus) have been consistently associated withthe topical ocular use of such compounds, in particular PGF_(2α) and itsprodrugs, e.g., its 1-isopropyl ester, in humans. The clinicalpotentials of prostaglandins in the management of conditions associatedwith increased ocular pressure, e.g. glaucoma are greatly limited bythese side effects.

U.S. Pat. No. 5,688,819, commonly assigned to Allergan, Inc., andincorporated herein by reference discloses compounds known asprostamides. Prostamides are distinguished from prostaglandins in thatthe oxygen which is bonded to carbonyl group is replaced by a nitrogenbearing substituent. Those skilled in the art will readily recognizethat this replacement significantly alters several electronic and stericproperties of an important structural feature in the biologicalmolecule. Significantly, it is commonly believed in the art thatresonance between the nitrogen lone pair and the carbonyl π-bond issignificantly greater than resonance between the carbonyl group and anoxygen lone pair in a carboxylic ester or a carboxylic acid. This beliefis supported by the well established experimental observation that thenitrogen atom in an amide is planar, as opposed to the pyramidalgeometry of an amine. Thus, the commonly accepted belief in the art isthat the nitrogen atom of an amine is sp³ hybridized, while nitrogenatom of an amide is sp² hybridized, with the bonded electrons occupyingthe sp² hybrid orbitals and the nonbonded electron pair occupying a porbital to allow for conjugation with the carbonyl π system. Bycontrast, the hybridization, bonding, and geometry of the electrons ofthe oxygen atom in water and alcohols are very similar to those ofcarboxylic acids or carboxylic esters.

The increased resonance between the nitrogen and the carbonyl group inthe amide confers several unique properties to the molecule. First, itis well known in the art that hydrolysis of amides is at least twoorders of magnitude slower than the hydrolysis of esters (see, forexample, Francis A. Carey, Organic Chemistry, New York: McGraw-Hill BookCompany, 1987, p. 779). Thus, hydrolysis of amides in vivo is slowed tosuch an extent that a prostamide cannot be considered to be a prodrug ofa prostaglandin. Second, the increased resonance significantly increasesthe barrier to rotation about the nitrogen-carbonyl sigma bond relativeto the analogous rotational barrier associated with esters andcarboxylic acids. Thus, a prostamide has a sterically significant,stable, rigid group replacing the oxygen atom of the prostaglandin. Thissignificant steric difference will have a significant effect in bindingto a number of receptor sites since geometry is important for manyreceptor sites. Since the carboxylic acid group of a prostaglandin is apolar, ionizable, group, with four potential hydrogen bond receivingelectron pairs, and in the case of the protonated acid, one potentialhydrogen bond donor, it is reasonable for a person of ordinary skill inthe art to believe that this functional group will be important to thebinding of the molecule to a number of receptors. It follows thatchanging the resonance properties, the hybridization of the bonding andnonbonding electrons, the geometry of the nitrogen atom, the number ofavailable hydrogen bonding sites, and the electronegativity of the ofthe nitrogen relative to oxygen, will confer significantly differentbiological properties to prostamides relative to prostaglandins.

Recently, it is becoming more commonly accepted in the art that amideshave distinct properties over carboxylic acids. For example, it has beenshown that anandamide, a common amide of arachidonic acid, hassignificant biological activity that arachidonic acid does not. Otherwork has also been done to show that amides have distinct activity ascompared to carboxylic acid, which has caused some in the field toclassify fatty acid amides as “a new family of biologically activelipids” (Bezuglov, et. al., “Synethesis and Biological Evaluation ofNovel Amides of Polyunsaturated Fatty Acids with Dopamine”, Bioorganic &Medicinal Chemistry Letters 11 (2001), 447-449).

It has been shown that prostamides have pronounced effects on smoothmuscle and are potent ocular hypotensive agents. Additionally,prostamides cause significantly lower ocular surface hyperemia thanprostaglandins. One prostamide exemplary of the these effects isbimatoprost, which is marketed by Allergan, Inc. under the trade nameLumigan®, which has the structure shown in Formula I below.

However, although bimatoprost is associated with significantly lesshyperemia and other irritating side effects compared to certainprostaglandins, further improvement is still highly desirable.

Trefoil peptides, or trefoil factor family (TFF) peptides are a class ofpeptides which comprise a common structural motif, known as the trefoildomain, as part of their structure. The trefoil motif comprises about 20to about 60 amino acid residues (usually about 40) containing sixcysteine residues. The six cysteine residues form three disulfidebridges that complete three loops in the peptide chain so that theroughly 40 residues have a clover-like shape, known as the trefoildomain. TFF-peptides can have one or two trefoil domains per molecule,and may comprise additional amino acid residues which are not part ofthe trefoil domain. To date, three types of TFF-peptides have beenisolated from humans-TFF1 (also known as pS2), TFF2 (also known as SP),and TFF3 (also known as ITF). TFF1 and TFF3 peptides each contain onetrefoil domain, while TFF2 peptides contain two trefoil domains. TFF1and TFF2 peptides are both produced by mucus-producing cells of stomach,while TFF3 peptides are produced by goblet cells of small and largeintestine.

All three forms of TFF-peptides are known to be produced in epithelialcells around areas of damage to mucus membrane, suggesting that trefoilshave a role in healing injury, particularly to epithelial cells. It isbelieved that TFF-peptides assist healing by both stabilizing mucusmembrane at the injury site and by stimulating repair. It has been shownthat TFF-peptides noncovalently link mucin, thus influencing therheology (e.g. increases viscosity) of mucus gels. [Hauser F, Poulsom R,Chinery R, et al, Proc Natl Acad Sci USA, 1993, vol. 90, pp. 6961-6965;and Babyatsky M W, deBeaumont M, Thim L, Podolky D K, Gastroenterology,1996, vol. 110, pp. 489-497]. TFF-peptides also appear to be responsiblefor promoting the migration of epithelial cells to the site of injury,thus stimulating repair. [Göke M, et al, Experimental Cell Research,2001, vol 264, pp. 337-344; and Playford R J, Journal of the RoyalCollege of Physicians of London, vol 31, pp. 37-40].

Although there is still a great deal unknown about the role of TFFpeptides on the ocular surface, in the lacrimal gland, in the efferentpassages, and in surrounding tissue, it is believed that TFF-peptidesmay be present during healing and other related processes in the eye.Biosynthesis and storage TFF1 and TFF3 peptides, but not TFF2, is knownto occur in the human conjunctival epithelium [Langer G, et al, InvestOphthalmol Vis Sci, 1999, vol. 40, pp. 2220-2224], and in vitro studieshave shown that TFF2 and TFF3 peptides promote the migration of woundedcorneal epithelial-cells from rabbits [Göke M, et al, Experimental CellResearch, 2001, vol 264, pp. 337-344]. However, to the best of ourknowledge, no direct relationship has been unambiguously establishedbetween TFF-peptides and any pathological condition affecting the eye.

SUMMARY OF THE INVENTION

Disclosed herein are dosage forms and methods which comprise aprostaglandin or a prostamide and a trefoil factor family peptide.

DETAILED DESCRIPTION OF THE INVENTION

Disclosed herein are dosage forms which comprise a prostaglandin or aprostamide and a trefoil factor family peptide. Also disclosed aremethods of treating ocular or conjunctival hyperemia in a personcomprising administering topically to an eye of said person atherapeutically effective amount of a trefoil factor family peptide,wherein said person is being treated for glaucoma with aprostaglandin-related compound.

In relation to the methods disclosed herein, the individual propertiesof the prostaglandin-related compound and the trefoil factor familypeptide may be considerations in determining how the two areadministered. In certain embodiments the prostaglandin-related compoundand trefoil factor family are administered in a single composition. Inother embodiments, the prostaglandin-related compound and the trefoilfactor family peptide are administered separately. In other embodiments,the prostaglandin-related compound and the trefoil factor family peptideare administered simultaneously. In other embodiments, theprostaglandin-related compound and the trefoil factor family peptide areadministered at substantially different times. In other embodiments, theprostaglandin-related compound and the trefoil factor family peptide areadministered with equal frequency. In other embodiments, theprostaglandin-related compound is administered more frequently than thetrefoil factor family peptide. In other embodiments, theprostaglandin-related compound is administered less frequently than thetrefoil factor family peptide.

A “prostaglandin-related compound” is broadly defined as any compoundrelated to a prostaglandin by being a natural prostaglandin, aprostaglandin analog, a prostaglandin receptor agonist, a prostamide, ora pharmaceutically acceptable salt, or a prodrug of any of the previousclasses. Natural prostaglandins can be described as derivatives ofprostanoic acid which have the following structural formula:

In relation to the structure above and prostaglandin-related compounds,term “α chain” refers to the top chain which is formed by the carbonatoms referred to as 1-7 in the structure above. The term “ω chain”refers to the bottom chain which is formed by the carbon atoms referredto as 13-20 in the structure above. The ring formed by the carbon atomsreferred to as 8-12 will be referred to as the “cyclopentyl ring” hereinfor convenience. Natural prostaglandins are characterized by thepresence of functional groups or double bonds on their cyclopentyl ring,and by the presence or absence of a cis double bond between carbons 5and 6, and by the presence or absence of a trans double bond betweencarbons 13 and 14. Such nomenclature is well known in the art. However,while not desiring to limit the scope of the invention in any way, someimportant groups of natural prostaglandin compounds are prostaglandin E,prostaglandin F, and prostaglandin D. Prostaglandin E is characterizedby a carbonyl group at carbon 9 and a hydroxyl group at carbon 11 whichis in the alpha configuration. One prostaglandin E which is of interestherein is prostaglandin E₁, which has a single covalent bond betweencarbons 5 and 6 and a double covalent bond between carbons 13 and 14.Another prostaglandin E of interest herein is prostaglandin E₂, whichhas a double covalent bond between carbons 5 and 6 and a double covalentbond between carbons 13 and 14. Thus, the subscript designates thenumber of carbon-carbon double bonds found in the basic prostaglandinstructure.

The compounds known collectively as prostaglandin F are characterized bythe common features that both carbons 9 and 11 have hydroxyl groupsattached. Similar to prostaglandin E the OH is in the α-configurationfor carbon 11, but the configuration of the OH at carbon 9 is designatedby a subscript. Thus, prostaglandin F_(2α), which is of particularinterest herein, is a prostaglandin F which has the OH of carbon 9 inthe α-configuration, and similar to prostaglandin E₂, prostaglandinF_(2α) has two double covalent carbon-carbon bonds between carbons 5 and6 and carbons 13 and 14.

The compounds known collectively as prostaglandin D are characterized bythe common features that carbon 9 is CHOH, where the OH is in theα-configuration, and carbon 11 is C═O. Similar to the previous examples,one prostaglandin D of interest herein is designated prostaglandin D₂,which indicates that the compound has two double covalent carbon-carbonbonds between carbons 5 and 6 and carbons 13 and 14.

A “prostaglandin analog” as used herein refers to a compound havingcertain structural similarities to the natural prostaglandins. An analoghas all of the features of a natural prostaglandin related to thecyclopentyl ring, including stereochemistry, the α-hydroxyl group atC15, and the presence or absence of double bonds at carbons 5, 6, 13 and14, or reasonable equivalents of those features. A reasonable equivalentto a feature is a feature that a person of ordinary skill in the artwould reasonably consider as having a similar purpose, but might enhancethe properties of the compound. While not intending to limit the scopeof the invention in any way, in general, an atom or functional groupwhich is isovalent or isoelectronic with the atom or functional group itis replacing would be a reasonable equivalent. Thus, for example, anα-CHSH group is a reasonable equivalent for an α-CHOH group, and a C═Sgroup is a reasonable equivalent for a C═O group. Another type ofreasonable equivalent has different electronic properties but similarsteric properties to the group it is replacing. Thus, F is a reasonableequivalent for H and OCH₃ is a reasonable equivalent for OH.

Beyond the similarities for the cyclopentyl ring and the double bondsindicated, a prostaglandin analog will have an α-chain and an ω-chainwhich are attached to adjacent atoms on the cyclopentyl ring. Themeanings of the cyclopentyl ring and the α and ω chains forprostaglandin analogs are broader than those of the naturalprostaglandins. For a prostaglandin analog, the “cyclopentyl ring” is afive-membered ring consisting of three or more carbon atoms, the“α-chain” has between 4 and 12 carbon atoms and the “ω-chain” hasbetween 4 and 20 carbon atoms. Either chain may comprise double ortriple covalent bonds, aromatic or aliphatic rings, and heteroatoms suchas S, O, N, and halogens. The only stereochemical requirements ofprostaglandin analogs are the same as those of the naturalprostaglandins they are associated with. Thus, for a prostaglandin Eanalog, carbon 9 and carbon 11 should be CHOH with the OH in theα-configuration, and the α- and ω-chains should have the α and βconfigurations respectively with relation to the connection to thecyclopentyl ring. The table below lists features which would be presentin analogs of several types of natural prostaglandins. Alternatively, areasonable equivalent for each feature might be present in the givenprostaglandin analog.

Prosta- glandin Analog C9 C11 C15 C5–C6 C13–C14 E C═O CH(OH) CH(OH) NANA α conf α conf E₁ C═O CH(OH) CH(OH) single bond trans double α conf αconf bond E₂ C═O CH(OH) CH(OH) cis double trans double α conf α confbond bond F CH(OH) CH(OH) CH(OH) NA NA α conf α conf F_(2α) CH(OH)CH(OH) CH(OH) cis double trans double α conf α conf α conf bond bond DCH(OH) C═O CH(OH) NA NA α conf α conf D₂ CH(OH) C═O CH(OH) cis doubletrans double α conf α conf bond bond NA means there is no requirement.

“A prostaglandin receptor agonist” refers to a compound which binds toand activates one of the prostaglandin receptors at a concentration ofless than 10⁴ nanomolar according to the Radioligand Binding and theFLIPR™ assay described hereafter. Of particular interest herein arecompounds having agonist activity at an FP receptor, an EP₂ receptor, anEP₄ receptor, and/or a DP receptor.

Radioligand Binding

Cells Stably Expressing EP₁, EP₂, EP₄ and FP Receptors

HEK-293 cells stably expressing the human or feline FP receptor, or EP₁,EP₂, or EP₄ receptors were washed with TME buffer, scraped from thebottom of the flasks, and homogenized for 30 sec using a Brinkman PT10/35 polytron. TME buffer was added to achieve a final 40 ml volume inthe centrifuge tubes (the composition of TME is 100 mM TRIS base, 20 mMMgCl₂, 2M EDTA; 10N HCl is added to achieve a pH of 7.4).

The cell homogenate was centrifuged at 19000 r.p.m. for 20 min at 4° C.using a Beckman Ti-60 rotor. The resultant pellet was resuspended in TMEbuffer to give a final 1 mg/ml protein concentration, as determined byBiorad assay. Radioligand binding competition assays vs. [³H-]17-phenylPGF_(2α) (5 nM) were performed in a 100 μl volume for 60 min. Bindingreactions were started by adding plasma membrane fraction. The reactionwas terminated by the addition of 4 ml ice-cold TRIS-HCl buffer andrapid filtration through glass fiber GF/B filters using a Brandel cellharvester. The filters were washed 3 times with ice-cold buffer and ovendried for one hour.

[³H-] PGE₂ (specific activity 180 Ci mmol) was used as the radioligandfor EP receptors. [³H] 17-phenyl PGF_(2α) was employed for FP receptorbinding studies. Binding studies employing EP₁, EP₂, EP₄ and FPreceptors were performed in duplicate in at least three separateexperiments. A 200 μl assay volume was used. Incubations were for 60 minat 25° C. and were terminated by the addition of 4 ml of ice-cold 50 mMTRIS-HCl, followed by rapid filtration through Whatman GF/B filters andthree additional 4 ml washes in a cell harvester (Brandel). Competitionstudies were performed using a final concentration of 5 nM [³H]-PGE₂, or5 nM [³H] 17-phenyl PGF_(2α) and non-specific binding determined with10⁻⁵M of unlabeled PGE₂, or 17-phenyl PGF_(2α), according to receptorsubtype studied.

Methods for FLIPR™ Studies

(a) Cell Culture

HEK-293(EBNA) cells, stably expressing one type or subtype ofrecombinant human prostaglandin receptors (prostaglandin receptorsexpressed: hDP/Gqs5; hEP₁; hEP₂/Gqs5; hEP_(3A)/Gqi5; hEP₄/Gqs5; hFP;hIP; hTP), were cultured in 100 mm culture dishes in high-glucose DMEMmedium containing 10% fetal bovine serum, 2 mM 1-glutamine, 250 μg/mlgeneticin (G418) and 200 μg/ml hygromycin B as selection markers, and100 units/ml penicillin G, 100 μg/ml streptomycin and 0.25 μg/mlamphotericin B.

(b) Calcium Signal Studies on the FLIPR™

Cells were seeded at a density of 5×10⁴ cells per well in Biocoat®Poly-D-lysine-coated black-wall, clear-bottom 96-well plates(Becton-Dickinson) and allowed to attach overnight in an incubator at37° C. Cells were then washed two times with HBSS-HEPES buffer (HanksBalanced Salt Solution without bicarbonate and phenol red, 20 mM HEPES,pH 7.4) using a Denley Cellwash plate washer (Labsystems). After 45minutes of dye-loading in the dark, using the calcium-sensitive dyeFluo-4 AM at a final concentration of 2 μM, plates were washed fourtimes with HBSS-HEPES buffer to remove excess dye leaving 100 μl in eachwell. Plates were re-equilibrated to 37° C. for a few minutes.

Cells were excited with an Argon laser at 488 nm, and emission wasmeasured through a 510-570 nm bandwidth emission filter (FLIPR™,Molecular Devices, Sunnyvale, Calif.). Drug solution was added in a 50μl volume to each well to give the desired final concentration. The peakincrease in fluorescence intensity was recorded for each well. On eachplate, four wells each served as negative (HBSS-HEPES buffer) andpositive controls (standard agonists: BW245C (hDP); PGE₂ (hEP₁;hEP₂/Gqs5; hEP_(3A)/Gqi5; hEP₄/Gqs5); PGF_(2α) (hFP); carbacyclin (hIP);U46619 (hTP), depending on receptor). The peak fluorescence change ineach drug-containing well was then expressed relative to the controls.

Compounds were tested in a high-throughput (HTS) orconcentration-response (CoRe) format. In the HTS format, forty-fourcompounds per plate were examined in duplicates at a concentration of10⁻⁵ M. To generate concentration-response curves, four compounds perplate were tested in duplicates in a concentration range between 10⁻⁵and 10⁻¹¹ M. The duplicate values were averaged. In either, HTS or CoReformat each compound was tested on at least 3 separate plates usingcells from different passages to give an n≧3.

Prostamides are also considered to be “prostaglandin-related” compounds.For the purposes of this disclosure, the term amide has the broadestmeaning generally understood by organic chemists. Prostamides areprepared by methods generally known in the art, and also by the methodsdescribed in U.S. Pat. No. 5,688,819, incorporated herein by reference.One important embodiment relates to the use of bimatoprost in thecompositions and methods disclosed herein. Bimatoprost is marketed underthe tradename Lumigan® by Allergan, Inc.

The term “prodrug” used herein has the meaning normally understood inthe art. That is, the prodrug is a compound which readily decomposes invivo to form a natural prostaglandin, a prostaglandin analog, aprostamide or a prostaglandin receptor agonist. While not intending tolimit the scope of the invention in any way, one common type of prodrugis an ester which hydrolyzes to yield an active compound with ahydroxide functional group.

The term “salt” has the meaning normally understood by those of ordinaryskill in the art. A “pharmaceutically acceptable salt” is any salt thatretains the activity of the parent compound and does not impart anydeleterious or untoward effect on the subject to which it isadministered and in the context in which it is administered.

Pharmaceutically acceptable salts of acidic functional groups may bederived from organic or inorganic bases. The salt may be a mono orpolyvalent ion. Of particular interest are the inorganic ions, lithium,sodium, potassium, calcium, and magnesium. Organic salts may be madewith amines, particularly ammonium salts such as mono-, di- and trialkylamines or ethanol amines. Salts may also be formed with caffeine,tromethamine and similar molecules. Hydrochloric acid or some otherpharmaceutically acceptable acid may form a salt with a compound thatincludes a basic group, such as an amine or a pyridine ring.

Certain terminology is used to refer to particular prostaglandin-relatedclasses of compounds. A compound which is referred to as “prostaglandinF-related” is a natural prostaglandin F, a prostaglandin F analog, aprostaglandin FP receptor agonist, or a prostamide having thecharacteristic features of prostaglandin F analog as describedpreviously, or a salt or a prodrug of any of the previous classes ofcompounds. Similar terminology can be used to identify otherprostaglandin compounds related to different classes of prostaglandinssuch as prostaglandin E (“prostaglandin E-related”) or prostaglandin D(“prostaglandin D-related”).

The quantity or concentration of a prostaglandin-related compound to beused in the compositions and methods disclosed herein can be determinedby one of ordinary skill in the art without undue experimentation. Inone embodiment, the concentration of the prostaglandin-related compoundin the dosage form in which it is administered is from 0.001% to 0.1%.In another embodiment, the concentration of the prostaglandin-relatedcompound in the dosage form in which it is administered is about 0.03%.

The term trefoil factor family (TFF) peptide as used herein refers toany peptide, whether natural or synthetic, which comprises the trefoilmotif described previously herein. That is, the TFF-peptide comprises aresidue comprising from 20 to about 60 amino acids, including sixcysteine residues. The cysteine residues form disulfide bonds whichcause the peptide residue to have a clover-like shape comprising threeloops. The methods of preparing of TFF-peptides, such as recombinantexpression of peptides and synthetic peptide synthesis, are well knownin the art. For example, methods of preparing TFF-peptides are includedin the following references: U.S. Pat. No. 6,525,018; Allen, et. al., JClin Gastroenterol 1998; 10 (Suppl 1): S93-S98; Ligumsky, et. al., Isr JMed Sci 1986; 22:801-806; Dignass, et. al., J. Clin. Invest., 94,376-383; Babyatsky, et. al., Gastroenterology, 110, 489-497; Hauser, et.al., Proc. Natl. Acad. Sci. USA, vol. 90, pp. 6961-6965, August 1993; WO02102403; and WO02085402, incorporated herein by reference. In oneembodiment the trefoil factor family peptide is TFF1, TFF2, or TFF3. Inanother embodiment the trefoil factor family peptide is TFF1 or TFF3.

The concentration or amount of the trefoil factor family peptide used inthe methods and compositions disclosed herein can readily be determinedby one of ordinary skill in the art without undue experimentation. Inone embodiment, the concentration of the trefoil factor family peptideis from 0.001% to 1%. In another embodiment, the concentration of thetrefoil factor family peptide is from 0.01% to 0.5%. In anotherembodiment, the concentration of the trefoil factor family peptide isfrom 0.1% to 0.2%. In another embodiment, the concentration of thetrefoil factor family peptide is about 0.15%.

A mucoadhesive is used in certain of the compositions and methodsdisclosed herein. With respect to this invention, the term“mucoadhesive” means a natural or synthetic component, includingmacromolecules, polymers, and oligomers, or mixtures thereof, that canadhere to a subject's mucous membrane. Adhesion of mucoadhesives to themucous membrane occurs primarily through noncovalent interactions, suchas hydrogen bonding and Van der Waal forces (Tabor et al., 1977 J.Colloid Interface Sci. 58:2 and Good 1977 J. Colloid Interface Sci.59:398). Examples of mucoadhesives for use in the embodiments disclosedherein include, but are not limited to, Carbopol®, pectin, alginic acid,alginate, chitosan, hyaluronic acid, polysorbates, such aspolysorbate-20, -21, -40, -60, -61, -65, -80, -81, -85;poly(ethyleneglycol), such as PEG-7, -14, -16, -18, -55, -90, -100,-135, -180, -4, -240, -6, -8, -9, -10, -12, -20, or -32;oligosaccharides and polysaccharides, such as Tamarind seedpolysaccharide, gellan, carrageenan, xanthan gum, gum Arabic, anddextran; cellulose esters and cellulose ethers; modified cellulosepolymers, such as carboxymethylcellulose, hydroxyethylcellulose,hydroxypropyl methylcellulose, hydroxyethyl ethylcellulose; polyetherpolymers and oligomers, such as polyoxyethylene; condensation productsof poly(ethyleneoxide) with various reactive hydrogen containingcompounds having long hydrophobic chains (e.g. aliphatic chains of about12 to 20 carbon atoms), for example, condensation products ofpoly(ethylene oxide) with fatty acids, fatty alcohols, fatty amides,polyhydric alcohols; polyether compounds, such as poly(methyl vinylether), polyoxypropylene of less than 10 repeating units; polyethercompounds, such as block copolymers of ethylene oxide and propyleneoxide; mixtures of block copolymers of ethylene oxide and propyleneoxide with other excipients, for example poly(vinyl alcohol);polyacrylamide; hydrolyzed polyacrylamide; poly(vinyl pyrrolidone);poly(methacrylic acid); poly(acrylic acid) or crosslinked polyacrylicacid, such as Carbomer®, i.e., a homopolymer of acrylic acid crosslinkedwith either an allyl ether of pentaerythritol, an allyl ether ofsucrose, or an allyl ether of propylene. In certain embodiments themucoadhesive is a polysaccharide. One polysaccharide which isparticularly useful as a mucoadhesive in the embodiments disclosedherein is Tamarind seed polysaccharide, which is a galactoxyloglucanthat is extracted from the seed kernel of Tamarindus Indica, and can bepurchased from TCI America of Portland, Oreg.

In certain embodiments a buffer is included to maintain the pH fromabout 6 to about 8. In particular cases, it is desirable to maintain thepH about 7. Buffers used are those known to those skilled in the art,and, while not intending to be limiting, some examples are acetate,borate, carbonate, citrate, and phosphate buffers. Preferably, thebuffer comprises borate. An effective amount of buffer necessary for thepurposes of this invention can be readily determined by a person skilledin the art without undue experimentation. In certain embodiments wherethe buffer comprises borate, the concentration of the borate buffer isabout 0.6%.

In any of the compositions described herein, a tonicity agent may beused. Tonicity agents are used in ophthalmic compositions to adjust theconcentration of dissolved material to the desired isotonic range.Tonicity agents are known to those skilled in the ophthalmic art, and,while not intending to be limiting, some examples include glycerin,mannitol, sorbitol, sodium chloride, and other electrolytes. Onparticularly useful tonicity agent is sodium chloride.

In any of the compositions are described herein, a preservative may beused, particularly when the composition is intended for multiple use.There may also be reasons to use a preservative in single usecompositions depending on the individual circumstances. The termpreservative has the meaning commonly understood in the ophthalmic art.Preservatives are used to prevent bacterial contamination inmultiple-use ophthalmic preparations, and, while not intending to belimiting, examples include benzalkonium chloride, stabilized oxychlorocomplexes (otherwise known as Purite®), phenylmercuric acetate,chlorobutanol, benzyl alcohol, parabens, and thimerosal. Oneparticularly useful preservative is benzalkonium chloride (BAK).

Under certain circumstances, a surfactant might be used in any of thecompositions related to this invention which are described herein. Theterm surfactant used herein has the meaning commonly understood in theart. Surfactants are used to help solubilize the therapeutically activeagent or other insoluble components of the composition, and may serveother purposes as well. Anionic, cationic, amphoteric, zwitterionic, andnonionic surfactants may all be used in this invention. Nonionicsurfactants, such as polysorbates, poloxamers, alcohol ethoxylates,ethylene glycol-propylene glycol block copolymers, fatty acid amides,alkylphenol ethoxylates, or phospholipids, are particularly useful forthe compositions and methods disclosed herein.

Another type of compound that might be used in any composition describedherein is a chelating agent. The term chelating agent refers to acompound that is capable of complexing a metal, as understood by thoseof ordinary skill in the chemical art. Chelating agents are used inophthalmic compositions to enhance preservative effectiveness. While notintending to be limiting, some useful chelating agents are edetatesalts, like edetate disodium, edetate calcium disodium, edetate sodium,edetate trisodium, and edetate dipotassium.

One particularly useful embodiment comprises a prostaglandin-relatedcompound at a concentration from 0.001% to 0.1%, a trefoil factor familypeptide, tamarind seed polysaccharide, about 0.5% sodium chloride, about0.005% benzalkonium chloride, and about 0.6% of a borate buffer whereinthe pH of the composition is adjusted to from about 6 to about 8. In onecomposition, the prostaglandin-related compound is bimatoprost, which ispresent at a concentration of 0.03%. In another composition, theprostaglandin-related compound is latanoprost, which is present at aconcentration of 0.005%. In another composition, theprostaglandin-related compound is travoprost, which is present at aconcentration of 0.004%. In another composition, theprostaglandin-related compound is unoprostone isopropyl, which ispresent at a concentration of 0.15%.

Another embodiment relates to a pharmaceutical product comprising acomposition comprising a therapeutically effective amount of aprostaglandin F-related compound and a therapeutically effectiveconcentration of a trefoil factor family peptide, and a package suitablefor ophthalmic use from which said composition is dispensed, wherein theuse of the composition for the prevention or treatment of glaucoma isindicated thereon.

The best mode of making and using the present invention are described inthe following examples. These examples are given only to providedirection and guidance in how to make and use the invention, and are notintended to limit the scope of the invention in any way.

EXAMPLE 1

Compositions related to this invention are prepared by the followingprocedure. Unless otherwise indicated, all procedural steps are carriedout at room temperature.

Part I

Tamarind seed polysaccharide (TSP) is added to purified water at theconcentration indicated in the Table 1, and the solution is brought to aboil and maintained at a gentle boil for about 30 minutes. The solutionis then allowed to cool to room temperature, and water is added tocompensate for evaporative loss during boiling. The solution is thenfiltered through a 20 micron clarity filter followed by a 0.45 micronsterilizing filter.

Part II

Each component listed in Table 1 is added in amount needed to providethe indicated concentration to a fixed volume of the solution from partI, in the following order: TFF 1, boric acid, sodium borate decahydrate,sodium chloride, and BAK. After the addition of each component, themixture is stirred until the solute is completely dissolved before thenext component is added. When all of the components of the formulationhave been added and dissolved, the pH is then adjusted to 7.0 with NaOHor HCl. The solution is then sterile filtered.

TABLE 1 Component Function % (w/v) Bimatoprost Prostamide 0.03 TFF 1TFF-peptide 0.15 Tamarind Seed Polysaccharide (TSP) Mucoadhesive 0.5Boric Acid Buffer 0.6 Sodium Borate Decahydrate Buffer 0.035 SodiumChloride Tonicity Agent 0.53 Benzalkonium Chloride (BAK) Preservative0.005 Purified Water Q.s. HCl or NaOH Buffer adjust to pH 7.0

EXAMPLE 2

A formulation having the composition of Table 2 is prepared according toan analogous procedure to that of Example 1. Latanoprost is well knownin the art, and can be prepared by procedures described in U.S. Pat. No.6,429,226, incorporated herein by reference.

TABLE 2 Component Function % (w/v) Latanoprost Prostaglandin 0.005 TFF 1TFF-peptide 0.15 Tamarind Seed Polysaccharide (TSP) Mucoadhesive 0.5Boric Acid Buffer 0.6 Sodium Borate Decahydrate Buffer 0.035 SodiumChloride Tonicity Agent 0.53 Benzalkonium Chloride (BAK) Preservative0.005 Purified Water Q.s. HCl or NaOH Buffer adjust to pH 7.0

EXAMPLE 3

A formulation having the composition of Table 3 is prepared according toan analogous procedure to that of Example 1.

TABLE 3 Component Function % (w/v) Bimatoprost Prostamide 0.03 TFF 1TFF-peptide 0.15 Sodium Carboxymethylcellulose Mucoadhesive 0.5 BoricAcid Buffer 0.6 Sodium Borate Decahydrate Buffer 0.035 Sodium ChlorideTonicity Agent 0.53 Benzalkonium Chloride (BAK) Preservative 0.005Purified Water Q.s. HCl or NaOH Buffer adjust to pH 7.0

EXAMPLE 4

A formulation having the composition of Table 4 is prepared according toan analogous procedure to that of Example 1.

TABLE 4 Component Function % (w/v) Bimatoprost Prostamide 0.03 TFF 3TFF-peptide 0.15 Hydroxypropylmethylcellulose Mucoadhesive 0.5 BoricAcid Buffer 0.6 Sodium Borate Decahydrate Buffer 0.035 Sodium ChlorideTonicity Agent 0.53 Benzalkonium Chloride (BAK) Preservative 0.005Purified Water Q.s. HCl or NaOH Buffer adjust to pH 7.0

EXAMPLE 5

A formulation having the composition of Table 5 is prepared according toan analogous procedure to that of Example 1.

TABLE 5 Component Function % (w/v) Bimatoprost Prostamide 0.03 BoricAcid Buffer 0.6 Sodium Borate Decahydrate Buffer 0.035 Sodium ChlorideTonicity Agent 0.53 Benzalkonium Chloride (BAK) Preservative 0.005Purified Water Q.s. HCl or NaOH Buffer adjust to pH 7.0

EXAMPLE 6

A formulation having the composition of Table 6 is prepared according toan analogous procedure to that of Example 1.

TABLE 6 Component Function % (w/v) Travoprost Prostaglandin 0.004 TFF 1TFF-peptide 0.15 Sodium Carboxymethylcellulose Mucoadhesive 0.5 BoricAcid Buffer 0.6 Sodium Borate Decahydrate Buffer 0.035 Sodium ChlorideTonicity Agent 0.53 Benzalkonium Chloride (BAK) Preservative 0.005Purified Water Q.s. HCl or NaOH Buffer adjust to pH 7.0

EXAMPLE 7

A formulation having the composition of Table 7 is prepared according toan analogous procedure to that of Example 1.

TABLE 7 Component Function % (w/v) Unoprostone isopropyl Prostamide0.15% TFF 3 TFF-peptide 0.15 Hydroxypropylmethylcellulose Mucoadhesive0.5 Boric Acid Buffer 0.6 Sodium Borate Decahydrate Buffer 0.035 SodiumChloride Tonicity Agent 0.53 Benzalkonium Chloride (BAK) Preservative0.005 Purified Water Q.s. HCl or NaOH Buffer adjust to pH 7.0

EXAMPLE 8

A drop of a composition prepared according to one of Examples 1 and 3-5is added at least once a day to several patients suffering fromglaucoma. Reduction in intraocular pressure is observed for allpatients, but with reduced hyperemia observed in the patients receivingthe compositions of Examples 1, 3 and 4, which have a trefoil factorfamily peptide, relative to the patients receiving the composition ofExample 5.

1. A dosage form comprising a prostamide and a trefoil factor familypeptide wherein the prostamide is bimatoprost and the peptide isselected form the group consisting of TFF1 and TFF3.
 2. The dosage formof claim 1 wherein the concentration of the prostamide is from 0.001% to0.1%.
 3. The dosage form of claim 1 wherein the concentration of thetrefoil factor family peptide is from 0.001% to 1%.
 4. The dosage formof claim 1 wherein the concentration of the trefoil factor familypeptide is from 0.01% to 0.5%.
 5. The dosage form of claim 1 wherein theconcentration of the trefoil factor family peptide is from 0.1% to 0.2%.6. The dosage form of claim 1 wherein the concentration of the trefoilfactor family peptide is about 0.15%.
 7. The dosage form of claim 1which further comprises a mucoadhesive.
 8. The dosage form of claim 1which further comprises a polysaccharide.
 9. The dosage form of claim 1which further comprises Tamarind seed polysaccharide.
 10. The dosageform of claim 1 wherein the trefoil family factor peptide is TFF3. 11.The dosage form of claim 1 wherein the trefoil family factor peptide isTFF1.
 12. The dosage form of claim 2 which comprises Tamarind seedpolysaccharide, about 0.5% sodium chloride, about 0.005% benzalkoniumchloride, and about 0.6% of a borate buffer wherein the pH of thecomposition is adjusted to from about 6 to about
 8. 13. The dosage formof claim 12 which comprises about 0.03% bimatoprost.
 14. Apharmaceutical composition for treatment of glaucoma comprising acomposition comprising a therapeutically effective amount of prostamideand a therapeutically effective amount of trefoil factor family peptidewherein the prostamide is bimatoprost and the peptide is selected fromthe group consisting of TFF1 and TFF3.